We propose to study biotin holoenzyme synthesis in Saccharomyces cerevisiae. Our primary objective is to elucidate the recognition mechanism which allows a single holoenzyme synthetase to incorporate biotin into many different biotin-requiring enzymes. We will use two experiment approaches: 1) The primary amino acid sequence will be determined for biotin containing peptides from two biotin-requiring enzymes of Saccharomyces - pyruvate carboxylase and urea amidolyase. 2) Mutants of Saccharomyces will be selected which lack activity for all of the biotin requiring enzymes. These mutants will be analyzed to determine whether a single mutation is responsible for the observed phenotype and whether the biochemical lesion occurs in the synthesis of a single non-specific biotin-holoenzyme synthetase. BIBLIOGRAPHIC REFERENCES: Roon, R.J., Even, H.L., Larimore, F. and Dunlop, P.C., "Methylamine and Ammonia Transport in Saccharomyces cerevisiae", J. Bacteriol. 122, 502 (1975). Dunlop, P.C., Roon, R.J. and Even, H.L., "Utilization of D-Asparagine by Saccharomyces cerevisiae", J. Bacteriol. In Press, March (1976).